Effects of stream environmental variables on the distribution and abundance of Ephemeroptera, Plecoptera and Trichoptera (EPT) assemblages in the headwaters of rivers and streams of Mongolia were investigated in this broad scale survey aimed to provide a baseline for future research and management of the country’s aquatic ecosystems. The survey was carried out in the first-, second- and third-order streams of 80 rivers selected from the three main watersheds of Mongolia, namely the Arctic Ocean Basin (AOB), Pacific Ocean Basin (POB) and Central Asian Internal Basin (CAIB). We sampled larvae of EPT by kick and D-nets from the headwater sites during summers of 2007–2009. Chemical and hydrological variables of the sites were measured, and habitat of those sites was assessed. We recorded 89 taxa belonging to 34 families of EPT orders during the survey. The family Baetidae (Ephemeroptera) was the dominant taxon among all sites. Taxon richness was higher in the POB and AOB than CAIB, while total EPT abundance was higher in the CAIB. Significant differences in the community variables, such as taxon richness, Shannon’s diversity index and evenness of EPT assemblages were observed among the basins. Results of canonical correspondence analysis suggested that altitude, dissolved oxygen, order and width of streams were the most important factors explaining the variability in EPT larval distribution in the headwater streams in Mongolia.